Recurrent Airway Obstruction
Recurrent Airway Obstruction (RAO), a respiratory disease similar to human asthma, is one of the most commonly diagnosed conditions affecting the lung of older horses all around world. It is a type-1 immediate hypersensitivity allergic disease involving a series of events that begin with reaginic antibodies, mainly IgE. These antibodies bind with high affinity to Fc epsilon-RI (FcεRI) receptors via the Fc portion that are found on the membrane of mast cells and basophils. Once bound, the antibodies can cross-link with environmental allergens, resulting in mast cell degranulation leading to the production of histamine and other chemical mediators that act together to induce airway inflammation. At least one reaginic antibody against Faenia rectivirgula, a common gram-positive bacteria, was found to be associated with lung diseases in RAO-affected and unaffected horses, finding the antibody present in about 15% in RAO-affected horses (28).
Most evidence suggests that RAO is the result of a pulmonary hypersensitivity to inhaled antigens. Exposure of affected horses to hay dust, pollens, and mold spores leads to neutrophil accumulation in the lung and bronchospasm. The identification of allergen-specific IgE in bronchoalveolar lavage (BAL) fluid and sera of affected horses supports the involvement of a late phase, IgE-mediated, hypersensitivity reaction in the pathogenesis of equine RAO. The cytokine profiles of horse affected by RAO have been measured, and a modified Th2-like immune response was thought to be involved with elevated IL-4, IL-13 and IFNg levels (6,22).
CC10 in RAO
Clara Cell 10 kDa protein (CC10) is one of the members of a family of anti-inflammatory defense proteins produced predominantly in the airway epithelium of man and animals. In humans it is also called human uteroglobin (See U.S. Pat. Nos. 6,255,281 and 7,846,899). CC10 has been shown to inhibit the entry of eosinophils and neutrophils into the human airways during inflammation in the lung, and to inhibit enzymes that release inflammatory mediators (23). Some of the members of this family have been directly linked to protection from asthma, allergy and related inflammatory responses. The gene that encodes the protein was found to be down regulated during RAO in horses (34), and the production of the CC10 protein was also reported to be reduced in RAO horses (24). The amount of CC10 in the horse serum was measured using antibodies produced in rabbits using a synthetic 21 amino acid peptide from horse CC10 (EPSKPDADMKAATTQLKTLV; SEQ ID NO:2). This peptide corresponded to amino acids 47-66 of horse CC10 protein (protein accession no. NP 001075327.1) with two amino acid variations.
A hypo-allergic vaccine to an environmental antigen of Timothy grass pollen Profilins was developed by engineering its structure (35). The engineered protein was demonstrated to induce IgG's and inhibit allergic patients' IgE antibody binding to Profilins to a similar degree as those induced by immunization with the wild type. IgG's inhibited Profilin-induced basophil degranulation. Another environmental antigen, a novel Fel d1 cat allergy vaccine, was developed that induced strong IgG response, which abrogated the allergy response in mice (32). Allergen-induced systemic basophil degranulation was shown inhibited in an Fc gamma-RIIb-dependent manner (26). In addition, antibodies have been found in healthy horses; elevated IgG response were found in healthy horses to auto antigen (1). IgGb specific to KLH was found elevated in healthy versus control horses. An inverse correlation was reported between BCG vaccination and incidence of IgE mediated allergic diseases (18). BCG immunization was demonstrated to inhibit IgE production by B cells.
Receptors for the Fc fragment of immunoglobulin-G (Fc gamma-R's) are important molecules not only to mediate and control the effectors' functions of IgG antibodies, but they also control the autoimmunity-tolerance balance in the periphery. In humans, three different types of Fc gamma-R's, belonging to the immunoglobulin gene super-family have been identified via Fc gamma-RI (CD64), Fc gamma-RII (CD32) and Fc gamma-RIII (CD16). In humans, myeloid cells express both activating and inhibitory receptors of the Fc gamma-RII family. Fc gamma-RIIA mediates processes associated with cell activation, while Fc gamma-RIIB down regulates such signaling. Differences in affinity for IgG between activating and inhibitory Fc gamma-R can result in substantial local changes in their relative concentrations with important functional consequences. Human airway smooth muscle cells express IgG-Fc gamma-RIIb with the potential to suppress remodeling and immune-modulation (36). A wide range of inflammatory and autoimmune diseases, such as vasculitis, glomerulonephritis, and autoimmune hemolytic anemia, seems to be mediated, in part, by Fc gamma-R′s.
Human Asthma and Animal Models
Human asthma is a multifactorial disease, and is usually a chronic lung condition which can develop at any age and progress to a devastating disease. Asthma is classified as a hypersensitivity disease (5,30). An estimate shows that there are more than 150 million people with asthma worldwide, of which 20.3 million are in America. It is also important to note that the trend of deaths and hospitalization due to asthma is increasing in the industrialized countries of the world. Despite extensive research into the pathophysiology and treatment of asthma, high morbidity continues to threaten the human population. Because asthma can be induced by multiple stimuli, no single theory satisfactorily explains all types and cases, thus making a therapeutic regimen cumbersome.
It is a well-established fact that the salient feature of human asthma is the tendency of the affected airways to overreact to a wide variety of nonspecific stimuli, which has led to the use of bronchial provocation tests as a method to diagnose the condition (19). This hyperreactivity is characterized by hyperresponsiveness and hypersensitivity of airways to various inflammatory mediators (13). This condition can result from abnormal tissue reaction in the airways either due to intrinsic stimuli arising from a biochemical, neurological, or humoral physiological imbalance in the body or due to extrinsic stimuli arising from the environment. The changes in the environment are detected in the airway mucosa by sensory receptors, which are richly innervated by sensory (afferent) fibers. Various respirable stimuli, including charged particles, allergens, irritants, as well as heat, cold and acid, activate sensory receptors.
Animal Models of Human Asthma:
To examine various factors and mechanisms involved in pathogenesis of asthma, a number of mammalian animal models have been identified and used to study human airway hyperreactivity. For example, guinea pigs, which are not naturally susceptible to asthma, are used because of the ability to generate an experimentally-inducible model of asthma with indices similar to human asthma (14,15). Another animal model for asthma is mice which again are not naturally susceptible to asthma, but asthma can be induced experimentally. One advantage to mice is the ability to study the genetic aspects of the disease (12).
Equine Models of Human Asthma:
A major advantage to an equine model is that horses get asthma naturally like people. Horses are important for the similarity between the signs and symptoms of recurrent airway obstruction (RAO) to those of asthma, for the similarity in the nature of disease progression to human asthma, and for the ability in horses to induce or precipitate an “asthmatic” episode (7,33). Histological findings such as thickening of epithelial basement membrane, edema, inflammatory infiltrate in bronchial walls, oversized submucosal glands, hypertrophy of bronchial wall muscle, and emphysematous changes are observed in both human asthma and in equine RAO (6,7,33). One difference is that the cellular infiltrate in RAO is mostly neutrophils, whereas esosinophils are more common with human asthma. Recently, identification of persistent nuclear factor kappa-B (NF-KB) activity in asthma and RAO clearly suggests that equine RAO is a better animal model of human asthma than those of other mammalian species (8,20). Chronic obstructive pulmonary disease (COPD) in horses, commonly called heaves, and currently termed recurrent airway obstruction (RAO), is an airway hyperreactivity disease comparable to human asthma (7,33). Horses with heaves spontaneously develop acute bronchiolitis after exposure to antigens such as hay or pasture mold. Heaves is a more common condition in temperate climatic regions where horses are confined in stables during winter seasons and where they are house near stored hay. Another form of RAO seen in the southern regions of the U.S. is summer pasture-associated obstructive pulmonary disease (SPAOPD), caused by allowing horses to graze on lush pasture in the humid summer months where molds and pollens in the grass serve as antigen stimulation (4). Both forms of RAO share common signs and symptoms (4,6,11). The clinically abnormal condition of RAO is characterized by hyperreactivity of airways to antigens, inflammatory mediators and various other external stimuli. The condition also presents a decreased dynamic compliance of lung and increased airflow resistance to histamine- or methacholine-challenges, which are the common tests performed for diagnosing either human asthma or equine RAO. When horses are exposed to diverse stimuli such as allergens or respirable irritants in the environment, their airways react with a specific response that includes infiltration of inflammatory cells in the lungs, damage of airway epithelium, and inflammatory mediator release. This reaction primes the airways to respond excessively to a broader range of stimuli. Continued stimulation leads to airway hyperreactivity, which is characterized by hypersensitivity (increased sensitivity of airway smooth muscles) and hyperresponsiveness (increased magnitude of smooth muscle contractility) to diverse stimuli as observed in human asthma (3,6,7,11). Thus, RAO in horses shares several common features of human asthma, such as airway hyperreactivity, severe bronchoconstriction, increased mucus production, presence of airway inflammatory mediators in bronchoalveolar lavage fluids, infiltration of inflammatory cells, difficulty in breathing and exercise intolerance. Horses then have several advantages as an animal model for human asthma. Of course, the main disadvantages to using horses are size and cost to obtain and house.
U.S. Pat. Nos. 7,846,899 and 6,255,281 and U.S. Patent Application Publication No. 2005/0261180 describe the use of recombinant human CC10 (also referred to as human uteroglobin) and its use in various human inflammatory and respiratory disorders.
U.S. Patent Application Publication No. 2011/0240012 describes the use of recombinant human CC10 and methods to administer CC10 nasally, and its use in treatment of various nasal inflammatory diseases, including rhinitis and sinusitis.